Global Wnt Inhibition With a Porcupine Inhibitor Decreases Established Trabecular Bone in a Mouse Model of Fibrous Dysplasia

Abstract

Background: G protein-coupled receptors (GPCRs) mediate a wide spectrum of physiological functions, including bone development and remodeling. Fibrous dysplasia (FD) is a common skeletal dysplasia caused by increased Gs-GPCR signaling and characterized by fibrotic expansile bone lesions. FD has no effective medical treatments. Our prior studies used the ColI(2.3)+/Rs1+ mouse model where osteoblastic-cell Gs signaling induced a dramatic FD-like phenotype and increased Wnt signaling, which we hypothesized is a major driver of the phenotype. Furthermore, we previously showed that blocking Rs1 expression could reverse the abnormal bone phenotype, providing proof-of-concept for finding therapies for FD. Methods: Long bone stromal cells from wildtype and ColI(2.3)+/Rs1+ 9-week-old male mice were analyzed by single-cell RNAseq to identify potential cellular sources of Wnt ligands. We compared these findings with global inhibition of Wnt activity by oral administration of the porcupine inhibitor LGK974 to ColI(2.3)+/Rs1+ mice. These mice were analyzed by histology and micro-computed tomography (micro-CT). Results: Control and ColI(2.3)+/Rs1+ bones showed similar scRNAseq results, except for a large expansion of osteoblastic lineage cells and increased differential expression (DE) genes in this cluster. Expression of Gi-GPCRs was increased, potentially as compensation for the strong Gs-GPCR pathway activation induced by Rs1 expression. We also found increased GH/IGF1 pathway activation in the osteoblastic cluster, and expression of multiple Wnt ligands within multiple cell clusters. We also identified a cell population unique to the ColI(2.3)+/Rs1+ FD-like bone lesions. Broad Wnt production inhibition of porcupine by LGK974 induced dose-dependent resorption of the abnormal FD bone shown by decreased bone volume and trabecular thickness; however, the fibrocellular infiltrate in the ColI(2.3)+/Rs1+ mice was still present. Conclusions: FD-like bones of ColI(2.3)+/Rs1+ mice showed broad activation of Wnt signaling in multiple cell types, suggesting both cell autonomous and non-cell autonomous activity. Broad Wnt inhibition decreased established FD-like trabecular bone, but the fibrocellular infiltrate did not fully reverse. These results suggest distinct roles of Gs-GPCR and Wnt signaling in FD pathogenesis.